Identification and characterization of molecules controlling embrionic stem cell (esc) differentiation

During the past decade Embryonic Stem cells (ESCs) became particularly attractive for translational medicine for their ability to give raise to specialized and functionally active cell types. However, before using ESCs for therapeutic purposes, such as transplantation and tissue regeneration, it will be essential to improve our ability to modulate and control ESC differentiation into lineage-specific derivatives. In this context, our main aim is to identify and characterize molecules able to control embryonic stem cell differentiation. To achieve this goal we have developed an in vitro system based on the RNA interference technology. By using this approach we have identified many factors strictly required for proper neuronal differentiation. Among them we have found a membrane protein that we have named Dies1. Dies1 suppression blocks ESC differentiation by maintaining the cells in an undifferentiated state. We have demonstrated that this protein is required to allow neuronal differentiation by participating to the signaling pathway of BMP4 that plays a crucial role in the balance between ESC stemness and differentiation. Indeed, Dies1 suppression induces an impairment of BMP4 signalling and on the other hand it induces an up-regulation of Nodal/Activin targets. Thus, Dies1 seems to be responsible for the balance between BMP4 and Nodal pathways in the decision of ESC fate. We have demonstrated a direct interaction between BMP4 receptor complex and Dies1 by means of site-direct mutagenesis followed by FRET microscopy and biochemical approaches. Moreover, we are characterizing the regulation of Dies1 in ESCs.